Neutrophils kill bacteria by means of a series of powerful oxidizing agents that they manufacture when they are in contact with their targets. The importance of these oxidizing agents in neutrophil function is exemplified by the clinical features of chronic granulomatous disease, an inherited disorder in which the inability of these cells to manufacture oxidizing agents is associated with a marked susceptibility to severe bacterial infections. The key enzyme responsible for the production of these oxidants is a membrane-bound NADPH oxidase that is dormant in resting cells but undergoes activation to catalyze the production of 02-from oxygen and NADPH when the cells encounter a stimulus such as opsonized bacteria. The proposals in this application are concerned with the characterization of this enzyme. The active oxidase will be studied using biochemical techniques to identify its components and their cofactors and to understand its mechanism of action. The resting enzyme will be purified, assaying with antibodies or by measuring its ability to undergo activation. Monoclonal antibodies will be prepared against the oxidase and used to study the enzyme from a variety of points of view. A cDNA for the oxidase will be cloned and used as a probe to study the properties of the oxidase gene.